First Morphological and Molecular Report of Aphelenchoides blastophthorus on Strawberry Plants in Switzerland.

Foliar nematodes represent a minor feeding group within the genus Aphelenchoides Fischer, 1894. The facultative plant parasitic species A. blastophthorus can cause crinkling of leaves, reduced vigor, and stunting of agricultural and ornamental plants. Here we report the first finding of A. blastophthorus in leaves, crowns, and roots of strawberry plants collected in Switzerland in 2018. Species identification was confirmed by morphological and morphometric characterization supported by molecular barcoding of 18S ribosomal RNA (18S), 28S ribosomal RNA (28S), and cytochrome c oxidase I (COI) gene fragment analyses. Phylogenetic analysis of 18S indicated that A. blastophthorus was grouped within close distance to A. fragariae, a well-known foliar nematode affecting strawberry plants. Furthermore, the newly generated molecular barcodes of the partial 28S and COI of A. blastophthorus will support species identification in the future.

Focused Metabolism of ß-Glucans by the Soil Bacteroidetes Species Chitinophaga pinensis.

The genome and natural habitat of Chitinophaga pinensis suggest it has the ability to degrade a wide variety of carbohydrate-based biomass. Complementing our earlier investigations into the hydrolysis of some plant polysaccharides, we now show that C. pinensis can grow directly on spruce wood and on the fungal fruiting body. Growth was stronger on fungal material, although secreted enzyme activity was high in both cases, and all biomass-induced secretomes showed a predominance of ß-glucanase activities. We therefore conducted a screen for growth on and hydrolysis of ß-glucans isolated from different sources. Most noncrystalline ß-glucans supported good growth, with variable efficiencies of polysaccharide deconstruction and oligosaccharide uptake, depending on the polysaccharide backbone linkage. In all cases, ß-glucan was the only type of polysaccharide that was effectively hydrolyzed by secreted enzymes. This contrasts with the secretion of enzymes with a broad range of activities observed during growth on complex heteroglycans. Our findings imply a role for C. pinensis in the turnover of multiple types of biomass and suggest that the species may have two metabolic modes: a "scavenging mode," where multiple different types of glycan may be degraded, and a more "focused mode" of ß-glucan metabolism. The significant accumulation of some types of ß-gluco-oligosaccharides in growth media may be due to the lack of an appropriate transport mechanism, and we propose that this is due to the specificity of expressed polysaccharide utilization loci. We present a hypothetical model for ß-glucan metabolism by C. pinensis that suggests the potential for nutrient sharing among the microbial litter community.IMPORTANCE It is well known that the forest litter layer is inhabited by a complex microbial community of bacteria and fungi. However, while the importance of fungi in the turnover of natural biomass is well established, the role of their bacterial counterparts is less extensively studied. We show that Chitinophaga pinensis, a prominent member of an important bacterial genus, is capable of using both plant and fungal biomass as a nutrient source but is particularly effective at deconstructing dead fungal material. The turnover of dead fungus is key in natural elemental cycles in the forest. We show that C. pinensis can perform extensive degradation of this material to support its own growth while also releasing sugars that may serve as nutrients for other microbial species. Our work adds detail to an increasingly complex picture of life among the environmental microbiota.

Chemical characterization and wound healing property of a ß-D-glucan from edible mushroom Piptoporus betulinus.

A water-soluble ß-D-glucan was obtained from fruiting bodies of Piptoporus betulinus, by hot aqueous extraction followed by freeze-thawing procedure and dialysis. Its molar mass distribution and conformational behavior in solution was assessed by size-exclusion chromatography coupled with multiangle laser light scattering, showing a polysaccharide with an average molecular weight of 2.5 × 105 Da with a random coil conformation for molecular weights below 1 × 106 Da. Typical signals of ß-(1 → 3)-linkages were observed in NMR spectrum (δ 102.7/4.76; 102.8/4.74; 102.9/4.52; and δ 85.1/3.78; 85.0/3.77) and also signals of O-6 substitution at δ 69.2/4.22 and 69.2/3.87. The analysis of partially O-methylated alditol acetates corroborates the NMR results, indicating the presence of a ß-D-glucan with a main chain (1 → 3)-linked, substituted at O-6 by single-units of glucose. The ß-D-glucan showed no toxicity on human colon carcinoma cell line (Caco-2) up to 1000 µg mL-1 and promoted cell migration on in vitro scratch assay, demonstrating a potential wound healing capacity.

Structure and antinociceptive effects of ß-D-glucans from Cookeina tricholoma.

Structurally different water-insoluble (1→3),(1→6) ß-D-glucans were isolated from aqueous and alkaline extracts of the mushroom-forming ascomycete Cookeina tricholoma, a wild edible mushroom found in Brazilian Amazon forest. The structures showed different substitution patterns, which may influence their extractability and consequently their conformation in solution, and different MW (4.3×10(5)Da, 3.7×10(5)Da and 8.2×10(5)Da, for ICW-Ct, IHW-Ct and IK2-Ct, respectively). The main-chains are composed of (1→3)-linked ß-D-Glcp units O-6 substituted by side chains with different lengths of (1→6)-linked ß-d-Glcp units (ICW-Ct and IHW-Ct) or by a combination of (1→6)-linked ß-D-Glcp units and single units of ß-D-Glcp (IK2-Ct). ß-D-glucans with similar MW and showing only (1→6)-linked ß-D-Glcp units as side chains (ICW-Ct and IHW-Ct) showed significant inhibition of neurogenic pain, 69±11 and 57±11% at the dose of 10mgkg(-1), respectively, in the model of nociception induced by intraplantar injection of formalin.

Mushroom heteropolysaccharides: A review on their sources, structure and biological effects.

Mushrooms have been largely studied not only by their d-glucans, but also because they present a class of more complex polymers: the heteropolysaccharides. Heteropolysaccharides show variability on their monosaccharide composition, anomeric configuration, linkage and branching type, besides some of these molecules can present natural methylated monosaccharides and also acid monosaccharides, which enhance the difficulty of the purification and characterization of their structure. As a result of such complexity, mushroom heteropolysaccharides can be considered an interesting source of molecules with medicinal and industrial applications. Consequently a plenty of new researches has been published in the past 12 years about the isolation, chemical characterization and biological activities of heteropolysaccharides from mushrooms, especially from Basidiomycetes. Therefore, this review intends to organize and classify the information described up to now about such polysaccharides obtained from different sources of mushroom-forming fungi, Basidiomycetes, Ascomycetes and Hybrid mushrooms, and provides a brief reflection on how the chemical studies have been carried out.

Glycan analysis of Fonsecaea monophora from clinical and environmental origins reveals different structural profile and human antigenic response.

Dematiaceous fungi constitute a large and heterogeneous group, characterized by having a dark pigment, the dihydroxynaftalen melanin-DHN, inside their cell walls. In nature they are found mainly as soil microbiota or decomposing organic matter, and are spread in tropical and subtropical regions. The fungus Fonsecaea monophora causes chromoblastomycosis in humans, and possesses essential mechanisms that may enhance pathogenicity, proliferation and dissemination inside the host. Glycoconjugates confer important properties to these pathogenic microorganisms. In this work, structural characterization of glycan structures present in two different strains of F. monophora MMHC82 and FE5p4, from clinical and environmental origins, respectively, was performed. Each one were grown on Minimal Medium (MM) and Czapeck-Dox (CD) medium, and the water soluble cell wall glycoconjugates and exopolysaccharides (EPS) were evaluated by NMR, methylation and principal component analysis (PCA). By combining the methylation and 2D NMR analyses, it was possible to visualize the glycosidic profiles of the complex carbohydrate mixtures. Significant differences were observed in ß-D-Galf-(1→5) and (1→6) linkages, α- and ß-D-Glcp-(1→3), (1→4), and (1→6) units, as well as in α-D-Manp. PCA from (1)H-NMR data showed that MMHC82 from CD medium showed a higher variation in the cell wall carbohydrates, mainly related to O-2 substituted ß-D-Galf (δ 106.0/5.23 and δ 105.3/5.23) units. In order to investigate the antigenic response of the glycoconjugates, these were screened against serum from chromoblastomycosis patients. The antigen which contained the cell wall of MMHC82 grown in MM had ß-D-Manp units that promoted higher antigenic response. The distribution of these fungal species in nature and the knowledge of how cell wall polysaccharides and glycoconjugates structure vary, may contribute to the better understanding and the elucidation of the pathology caused by this fungus.

Water-soluble polysaccharides from Pleurotus ostreatus var. florida mycelial biomass.

Pleurotus ostreatus var. florida known as Hiratake has a high nutritional value, presents medicinal and nutraceutical properties and it is one of the consumed mushrooms in Brazil. Thus, the aim of this study was to characterize the chemical structure of polysaccharides found in mycelial biomass produced by submerged culture of P. ostreatus var. florida in order to compare with those found in P. ostreatus var. florida fruit bodies. Aqueous and alkali extracts obtained from mycelial biomass were purified, 13C NMR, GC-MS and chemical techniques were used to characterize three polysaccharide structures: a mannogalactan (MG-PfM) with α-D-Galp and 3-O-Me-α-D-Galp units, both (1→6)-linked, highly substituted at O-2 by D-Manp, a glycogen-like polymer (GLY-PfM) with α-D-Glp (1→4)-linked main chain, partially substituted at O-6 by α-D-Glcp side chains and a (1→3), (1→6) ß-D-glucan (ßGLC-PfM) with a main chain of ß-D-Glcp (1→3)-linked units, partially substituted at O-6 by side chains of 6-O-substituted ß-D-glucopyranosyl units, on an average of one to every two residues of the backbone. These results show the possibility to obtain similar and also different molecules from those found in the fruiting body of the same mushroom species, therefore the submerged culture of mushroom is a promising way to give raise molecules of interest.

Polysaccharide glucomannan isolated from Heterodermia obscurata attenuates acute and chronic pain in mice.

Glucomannan (GM) is a polysaccharide obtained from Heterodermia obscurata lichens. The present study was conducted to elucidate the antinociceptive effect of GM in behavioural models of acute and chronic pain in mice. GM reduced mechanical allodynia and the levels of interleukin 1-ß (IL-1ß) in spinal cord and nerve in the partial sciatic nerve ligation (PSNL) model. Systemic treatment with GM inhibited the nociception induced by intraplantar injection of glutamate and by intrathecal injection of N-methyl-d-aspartic acid (NMDA), (±)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD), tumour necrosis factor α (TNF-α) and IL-1ß. Taken together, our data demonstrate that GM has significant antinociceptive effect in acute and chronic pain, suggesting a potential interest in the development of new clinically relevant drugs for the management of pain.

Agaricus bisporus fucogalactan: structural characterization and pharmacological approaches.

The fucogalactan from Agaricus bisporus (EFP-Ab) obtained on aqueous extraction followed by purification had M(w) 37.1 × 10(4)g mol(-1) relative to a (1→6)-linked α-D-Galp main-chain partially methylated at HO-3, and partially substituted at O-2 by nonreducing end-units of α-L-Fucp or ß-d-Galp. EFP-Ab also inhibited significantly the neurogenic and inflammatory phases of formalin-induced licking, however, the antinociceptive effect was more pronounced against the inflammatory phase with ID(50) of 36.0 (25.8-50.3)mg kg(-1). In addition, EFP-Ab decreased the lethality induced by CLP. Its administration reduced the late mortality rate by 40%, prevented neutrophil accumulation in lungs and markedly decreased iNOS and COX-2 protein expression by ileum cells. These data show for the first time that EFP-Ab has significant anti-sepsis, antinociceptive and anti-inflammatory actions, which seems to be related to the decreased iNOS and COX-2 expression. Collectively, the present results demonstrate that EFP-Ab could constitute an attractive molecule of interest for the development of new drugs.

Polysaccharides from Agaricus bisporus and Agaricus brasiliensis show similarities in their structures and their immunomodulatory effects on human monocytic THP-1 cells.

BACKGROUND: Mushroom polysaccharides have traditionally been used for the prevention and treatment of a multitude of disorders like infectious illnesses, cancers and various autoimmune diseases. Crude mushroom extracts have been tested without detailed chemical analyses of its polysaccharide content. For the present study we decided to chemically determine the carbohydrate composition of semi-purified extracts from 2 closely related and well known basidiomycete species, i.e. Agaricus bisporus and A. brasiliensis and to study their effects on the innate immune system, in particular on the in vitro induction of pro-inflammatory cytokines, using THP-1 cells. METHODS: Mushroom polysaccharide extracts were prepared by hot water extraction and precipitation with ethanol. Their composition was analyzed by GC-MS and NMR spectroscopy. PMA activated THP-1 cells were treated with the extracts under different conditions and the production of pro-inflammatory cytokines was evaluated by qPCR. RESULTS: Semi-purified polysaccharide extracts of A. bisporus and A. brasiliensis (= blazei) were found to contain (1→6),(1→4)-linked α-glucan, (1→6)-linked ß-glucan, and mannogalactan. Their proportions were determined by integration of 1H-NMR signs, and were considerably different for the two species. A. brasiliensis showed a higher content of ß-glucan, while A. bisporus presented mannogalactan as its main polysaccharide. The extracts induced a comparable increase of transcription of the pro-inflammatory cytokine genes IL-1ß and TNF-α as well as of COX-2 in PMA differentiated THP-1 cells. Pro-inflammatory effects of bacterial LPS in this assay could be reduced significantly by the simultaneous addition of A. brasiliensis extract. CONCLUSIONS: The polysaccharide preparations from the closely related species A. bisporus and A. brasiliensis show major differences in composition: A. bisporus shows high mannogalactan content whereas A. brasiliensis has mostly ß-glucan. Semi-purified polysaccharide extracts from both Agaricus species stimulated the production of pro-inflammatory cytokines and enzymes, while the polysaccharide extract of A. brasiliensis reduced synthesis of these cytokines induced by LPS, suggesting programmable immunomodulation.

Polysaccharides present in cultivated Teloschistes flavicans symbiosis: comparison with those of the thallus.

The chemical structures of polysaccharides present in aposymbiotically cultured myco- and photobionts of the lichen Teloschistes flavicans were determined, in order to compare them with those previously found in the intact thallus. The mycobiont was cultured on a solid Lilly and Barnett medium and the resulting colonies were freeze dried, defatted, and their polysaccharides were extracted successively with 2%, 10% and 30% aq. KOH, each at 100 degrees C. The extracts were neutralized (HOAc) and fractionated, giving rise to three homogeneous fractions, PFSK2 from 2% KOH, which contained a (1-->4),(1-->6)-linked alpha-glucan (1:1 ratio, pullulan), fraction PK10 from 10% KOH extraction, which was a linear (1-->3)-linked linear beta-glucan (laminaran), and fraction PK30 from 30% KOH extraction, being a branched (1-->3),(1-->6)-linked beta-glucan. The photobiont (Trebouxia sp. de Puymaly) was cultured in liquid nutrient medium, and after purification, a linear (1-->5)-linked beta-galactofuranan was characterized. The galactofuranan and the laminaran were not present in the symbiotic thallus, in contrast to the glucans, showing that the mycobiont alone produces them without participation of the photobiont.